Cystic ovarian disease in dairy cattle: Diagnostic accuracy when using B-mode and color Doppler ultrasound.

The most frequently reported definition of cystic ovarian disease in cattle is an abnormally persistent follicle (>7 to 10 d) with a diameter >25 mm. Discrimination between luteal and follicular ovarian cystic structures has traditionally been conducted by measuring the rim width of luteal tissue. The most common practice used in the field for diagnosis of cystic ovarian disease is examination by rectal palpation with or without the use of a B-mode ultrasound. Color Doppler ultrasound technology allows assessment of blood flow area measurements in the ovary, which has been proposed as a potential indirect measure for plasma progesterone (P4) concentrations. The objective of this study was to compare the diagnostic accuracy of differentiating luteal structures from follicular ovarian cysts using measures collected with B-mode and color Doppler transrectal ultrasonography. The definition of an ovarian cyst was a follicle greater than 20 mm in diameter in the absence of a corpus luteum that persisted for at least 10 d. A 3-mm luteal rim width was used to differentiate follicular and luteal cysts. A total of 36 cows were enrolled in the study during routine herd reproductive examination visits, with 26 and 10 having follicular and luteal cysts, respectively. Cows enrolled in the study were examined using a Mini-ExaPad mini ultrasound with color Doppler capabilities (IMV Imaging Ltd.). Blood samples were collected from each cow to measure P4 serum concentrations. History and signalment of each cow, including days in milk, lactation, times bred, days since last heat, milk composition, and somatic cell counts, were retrieved from an online database (DairyComp 305, Valley Agricultural Software). The accuracy of diagnosing follicular from luteal cysts based on luteal rim thickness was analyzed by receiver operating characteristic (ROC) curve using P4 as the gold standard, where P4 concentrations exceeding 1 ng/mL was defined as luteal, and all other structures with less P4 were considered follicular. Luteal rim and blood flow area were selected for further analysis because they presented the best ROC curves for differentiating cystic ovarian structures, with areas under the curve of 0.80 and 0.76, respectively. Luteal rim width of 3 mm was used as the cutoff standard in the study, resulting in sensitivity and specificity of 50% and 86%, respectively. Blood flow area of 0.19 cm2 was used as the cutoff standard in the study, resulting in sensitivity and specificity of 79% and 86%, respectively. When combining the use of luteal rim width and blood flow area to differentiate cystic ovarian structures, a parallel approach resulted in sensitivity and specificity of 73% and 93%, respectively, whereas an in-series approach resulted in sensitivity and specificity of 35% and 100%, respectively. In conclusion, the use of color Doppler ultrasonography when discriminating between luteal and follicular ovarian cysts in dairy cattle resulted in higher diagnostic accuracy compared with using B-mode ultrasonography alone.

A novel risk assessment tool for bovine respiratory disease in preweaned dairy calves.

Due to the increased morbidity and mortality of bovine respiratory disease (BRD) in dairy calves, as well as an increasing urgency for the judicious use of antimicrobials in farm animals, a comprehensive risk assessment tool for BRD in preweaned dairy calves has been designed based on a longitudinal and a cross-sectional study. As a multifactorial disease complex in which immune function stressors increase susceptibility to respiratory pathology, risk management programs for environmental and husbandry practices may be an effective approach for BRD control. Practices of known or suspected effect on BRD in preweaned calves have been explored in 2 large studies correlating management factors to BRD prevalence (BRD 100 study) and incidence (BRD 10K study) and forming the scores presented here. Priority was given to results from multivariable over univariable model estimates. However, when used, univariable model estimates were adjusted for confounders or stratified by effect modifiers if necessary. Regression coefficients were translated into scores, which are presented in a field-ready tool consisting of (1) a risk assessment questionnaire, which identifies the herd-specific risk factors and the risk scores associated with each; (2) the California BRD scoring system to estimate the BRD prevalence at the time of risk assessment for future comparison with the prevalence after interventions; and (3) the BRD control and prevention herd management plan, which can be used to plan and track the interventions identified. Scores for 100 dairies across California were used to benchmark a dairy's risk on a spectrum. With the help of the risk assessment tool, dairy producers, calf managers, and veterinarians may be able to adjust management factors that affect BRD risk on a farm and objectively monitor BRD prevalence before and after management interventions. As a result, the BRD risk assessment tool described here is the first comprehensive effort for herd-specific BRD control and prevention.

Management factors associated with bovine respiratory disease in preweaned calves on California dairies: The BRD 100 study.

The objective of this cross-sectional study was to determine how management practices on California dairies may be associated with bovine respiratory disease (BRD) in preweaned calves. A convenience sample of 100 dairies throughout California, providing a study population of 4,636 calves, were visited between May 2014 and April 2016. During each farm visit, in-person interviews with the herd manager or calf caretaker were conducted to collect information about herd demographics, maternity pen, colostrum and calf management, herd vaccinations, and dust abatement. A random sample of preweaned calves was identified and evaluated for the presence of BRD using a standardized tool. A survey-adjusted generalized linear mixed model with a logit link function was fitted with calf as the unit of analysis and dairy as the random effect. Mean study herd size (±SE) was 1,718 (±189.9) cows. Survey-adjusted estimates of breed types in the sample were 81.6% (±0.6) Holstein, 13.1% (±0.4) Jersey, and 5.3% (±0.5) crossbred or other purebred breeds, and calf sex proportions were 73.8% (±1.0) female and 26.2% (±1.0) male. Overall survey-adjusted BRD prevalence in the study herds was 6.91% (±0.69). Housing factors positively associated with BRD were metal hutches compared with wood hutches [odds ratio (OR) = 11.19; 95% confidence interval (CI) = 2.80-44.78], calf-to-calf contact in calves >75 d of age (OR = 9.95, 95% CI = 1.50-65.86), feeding Holstein calves <2.84 L of milk or replacer per day (OR = 7.16, 95% CI = 1.23-41.68), and lagoon water used for flushing manure under hutches compared with no flush (OR = 12.06, 95% CI = 1.93-75.47). Providing extra shade over hutches (OR = 0.08; 95% CI = 0.02-0.37), feeding calves at least 90% saleable milk (OR = 0.27, 95% CI = 0.13-0.54) or pasteurized milk (OR = 0.10; 95% CI = 0.03-0.36), and feeding >5.68 L of milk or replacer per day to Jersey calves (OR = 0.04; 95% CI = 0.01-0.28) were negatively associated with BRD. Our study identified management practices on California dairies with variability and that may contribute to differences in BRD prevalence, which will be incorporated into a risk-assessment tool to control and prevent BRD in preweaned dairy calves.

Effect of the environment on the risk of respiratory disease in preweaning dairy calves during summer months.

Heat stress has the potential to adversely affect the physiology, passive immunity, and growth of preweaning dairy calves, increasing their risk of respiratory disease. The effect of heat stress on the risk for bovine respiratory disease (BRD) may be mediated in part through housing, ventilation, and management factors. As a result, differences may exist in meteorological measures recorded in the calf-rearing area (macroenvironment) and within a calf's enclosure (microenvironment). The objective of this prospective cohort study was to evaluate and compare the association between exposure to temperature and humidity measured at the macro- and microenvironment, and BRD in preweaning dairy calves; a secondary objective was to evaluate the correlation between the macro- and microenvironment. A cohort of 252 calves from 4 premises in central San Joaquin Valley, California (CA), was followed and evaluated for development of respiratory disease using the CA BRD scoring system for preweaning dairy calves, a standardized and validated scoring system. During this time, the meteorological conditions of the calf-rearing area and the within-hutch environment were measured and showed a significant correlation with regard to temperature and humidity. Mixed effects logistic regression and survival analysis were used to analyze the association between the exposures daily environmental measures of temperature, humidity, and temperature-humidity index (THI) and the outcome BRD, adjusted for dairy premises, calf age, sex, and breed. Results showed a significant positive association between daily maximum temperature and BRD in both the calf's macroenvironment [odds ratio = 1.121 (95% confidence interval (CI) = 1.029-1.222)] and microenvironment [odds ratio = 1.203 (95% CI = 1.020-1.418)]. Estimated hazard rates also showed a significant positive association between BRD and daily maximum temperature in both the macroenvironment [hazard ratio = 1.127 (95% CI = 1.053-1.206)] and microenvironment [hazard ratio = 1.119 (95% CI = 1.047-1.197)]. In contrast, we found no association between daily maximum humidity in a calf's microenvironment and BRD. Daily maximum THI within the hutch was significantly associated with only the rate of BRD cases [hazard ratio = 1.070 (95% CI = 1.003-1,141)] but not the odds of occurrence of BRD. Maximum THI is estimated using temperature and humidity, which in California's hot and dry summers may limit variability in THI, explaining its weaker significant association with risk of BRD (or lack of association with odds of BRD) compared with models for maximum temperature in this study. Calves exposed to high day temperatures and relatively low humidity may be experiencing heat stress that predisposes to BRD. Results of the current study suggest that heat abatement efforts should address heat stress at the microenvironment level to mitigate BRD in calves. Further research should investigate strategies to improve calf hutch systems, including hutch materials and design that may optimize ventilation, provide ample shade, spacing, cleanliness, and protection from heat.

Pharmacokinetics of tulathromycin in plasma and milk samples after a single subcutaneous injection in lactating goats (Capra hircus).

Eight adult female dairy goats received one subcutaneous administration of tulathromycin at a dosage of 2.5 mg/kg body weight. Blood and milk samples were assayed for tulathromycin and the common fragment of tulathromycin, respectively, using liquid chromatography/mass spectrometry. Pharmacokinetic disposition of tulathromycin was analyzed by a noncompartmental approach. Mean plasma pharmacokinetic parameters (±SD) following single-dose administration of tulathromycin were as follows: C(max) (121.54 ± 19.01 ng/mL); T(max) (12 ± 12-24 h); area under the curve AUC(0→∞) (8324.54 ± 1706.56 ng·h/mL); terminal-phase rate constant λz (0.01 ± 0.002 h⁻¹); and terminal-phase rate constant half-life t1/2λz (67.20 h; harmonic). Mean milk pharmacokinetic parameters (±SD) following 45 days of sampling were as follows: Cmax (1594 ± 379.23 ng/mL); Tmax (12 ± 12-36 h); AUC(0→∞) (72,250.51 ± 18,909.57 ng·h/mL); λz (0.005 ± 0.001 h⁻¹); and t(1/2λz) (155.28 h; harmonic). All goats had injection-site reactions that diminished in size over time. The conclusions from this study were that tulathromycin residues are detectable in milk samples from adult goats for at least 45 days following subcutaneous administration, this therapeutic option should be reserved for cases where other treatment options have failed, and goat milk should be withheld from the human food chain for at least 45 days following tulathromycin administration.

Pharmacokinetics of ceftiofur crystalline free acid after single and multiple subcutaneous administrations in healthy alpacas (Vicugna pacos).

Six adult male alpacas received one subcutaneous administration of ceftiofur crystalline free acid (CCFA) at a dosage of 6.6 mg/kg. After a washout period, the same alpacas received three subcutaneous doses of 6.6 mg/kg CCFA at 5-day intervals. Blood samples collected from the jugular vein before and at multiple time points after each CCFA administration were assayed for ceftiofur- and desfuroylceftiofur-related metabolite concentrations using high-performance liquid chromatography. Pharmacokinetic disposition of CCFA was analyzed by a noncompartmental approach. Mean pharmacokinetic parameters (± SD) following single-dose administration of CCFA were Cmax (2.7 ± 0.9 µg/mL); Tmax (36 ± 0 h); area under the curve AUC0→∞ (199.2 ± 42.1 µg·h/mL); terminal phase rate constant λz (0.02 ± 0.003/h); and terminal phase rate constant half-life t1/2λz (44.7 h; harmonic). Mean terminal pharmacokinetic parameters (±SD) following three administrations of CCFA were Cmax (2.0 ± 0.4 µg/mL); Tmax (17.3 ± 16.3 h); AUC0→∞ (216.8 ± 84.5 µg·h/mL); λz (0.01 ± 0.003/h); and t1/2λz (65.9 h; harmonic). The terminal phase rate constant and the Tmax were significantly different between single and multiple administrations. Local reactions were noted in two alpacas following multiple CCFA administrations.

Short communication: Identification of coagulase-negative staphylococcus species from goat milk with the API Staph identification test and with transfer RNA-intergenic spacer PCR combined with capillary electrophoresis.

Coagulase-negative staphylococci (CNS) are the most commonly isolated bacteria from goat milk, but they have often been identified with phenotypic methods, which may have resulted in misclassification. The aims of this paper were to assess the amount of misclassification of a phenotypic test for identifying CNS species from goat milk compared with transfer RNA intergenic spacer PCR (tDNA-PCR) followed by capillary electrophoresis, and to apply the tDNA-PCR technique on different capillary electrophoresis equipment. Milk samples were collected from 416 does in 5 Californian dairy goat herds on 3 occasions during lactation. In total, 219 CNS isolates were identified at the species level with tDNA-PCR and subjected to the API 20 Staph identification test kit (API Staph; bioMérieux, Durham, NC). If the same species was isolated multiple times from the same udder gland, only the first isolate was used for further analyses, resulting in 115 unique CNS isolates. According to the tDNA-PCR test, the most prevalent CNS species were Staphylococcus epidermidis, Staphylococcus caprae, and Staphylococcus simulans. Typeability with API staph was low (72%). Although the API Staph test was capable of identifying the majority of Staph. epidermidis and Staph. caprae isolates, sensitivity for identification of Staph. simulans was low. The true positive fraction was high for the 3 most prevalent species. It was concluded that the overall performance of API Staph in differentiating CNS species from goat milk was moderate to low, mainly because of the low typeability, and that genotypic methods such as tDNA-PCR are preferred.

Progesterone milk residues in goats treated with CIDR-G(®) inserts.

Progesterone (P4)-impregnated intravaginal controlled internal drug-releasing devices (CIDRs) have been used worldwide for estrus synchronization in ruminants. CIDRs serve to place all treated animals in the luteal phase of the estrous cycle. The objectives of this study were to compare P4 concentrations in milk from normal reproductively cycling, CIDR-treated, and pregnant goats. CIDRs were placed in treatment goats on day 0 and removed on day 19. Milk was collected daily from day 0 to day 21 from control and CIDR-treated goats and for 5 consecutive days between 40 and 60 days of gestation from pregnant does. Milk P4 was plotted against time (in days) for each individual, and the area under the curve (AUC) was calculated as an estimate of total milk P4. The AUC(day 0-21) for control and CIDR-treated goats were 29.5 ± 11.9 and 33.7 ± 6.6 d·ng/mL, respectively (P = 0.77). The highest single-day and highest 5-day average P4 values for each animal were also compared among groups. Single-day peak P4 levels were 4.8 ± 1.5, 4.0 ± 1.0, and 6.0 ± 0.4 ng/mL for control, CIDR-treated, and pregnant goats (P = 0.42). The highest 5-day average P4 concentrations were 3.6 ± 1.3, 2.9 ± 1.8, and 4.2 ± 0.3 for control, CIDR-treated, and pregnant goats (P = 0.56). The results of this study show that intravaginal P4 CIDR devices inserted for 19 days in healthy goats resulted in milk P4 levels similar to or less than those endogenously produced during diestrus or pregnancy.

Production and processing of milk from transgenic goats expressing human lysozyme in the mammary gland.

The potential for applying biotechnology to benefit animal agriculture and food production has long been speculated. The addition of human milk components with intrinsic antimicrobial activity and positive charge to livestock milk by genetic engineering has the potential to benefit animal health, as well as food safety and production. We generated one line of transgenic goats as a model for the dairy cow designed to express human lysozyme in the mammary gland. Here we report the characterization of the milk from 5 transgenic females of this line expressing human lysozyme in their milk at 270 microg/mL or 68% of the level found in human milk. Milk from transgenic animals had a lower somatic cell count, but the overall component composition of the milk and milk production were not different from controls. Milk from transgenic animals had a shorter rennet clotting time and increased curd strength. Milk of such nature may be of benefit to the producer by influencing udder health and milk processing.

Accelerated regression of brain metastases in patients receiving whole brain radiation and the topoisomerase II inhibitor, lucanthone.

PURPOSE: To determine if lucanthone crossed the blood-brain barrier in experimental animals; and to determine accelerated tumor regression of human brain metastases treated jointly with lucanthone and whole brain radiation. METHODS AND MATERIALS: The organ distribution of 3H lucanthone in mice and 125I lucanthone in rats was determined to learn if lucanthone crossed the blood-brain barrier. Size determinations were made of patients' brain metastases from magnetic resonance images or by computed tomography before and after treatment with 30 Gy whole brain radiation alone or with lucanthone. RESULTS: The time course of lucanthone's distribution in brain was identical to that in muscle and heart after intraperitoneal or intravenous administration in experimental animals. Lucanthone, therefore, readily crossed the blood-brain barrier in experimental animals. CONCLUSION: Compared with radiation alone, the tumor regression in patients with brain metastases treated with lucanthone and radiation was accelerated, approaching significance using a permutation test at p = 0.0536.

A POLYGEN-adjuvanted killed Neospora caninum tachyzoite preparation failed to prevent foetal infection in pregnant cattle following i.v./i.m. experimental tachyzoite challenge.

Cattle immunised with a POLYGEN-adjuvanted killed Neospora caninum tachyzoite preparation were previously shown to produce interferon (IFN)-gamma at levels similar to those of tachyzoite-infected cattle. In view of the critical role of IFN-gamma in resistance of mice to N. caninum infection, these results prompted us to test the POLYGEN-adjuvanted preparation in pregnant cattle to determine whether it will be able to prevent foetal infection following an experimental tachyzoite challenge. Seven heifers were immunised at 35 and 63 days of gestation with the POLYGEN-adjuvanted preparation, while five heifers were inoculated with POLYGEN alone at the same days of gestation. Four weeks later, all heifers were challenged with a combined i.v./i.m. inoculation of tachyzoites. The same challenge was given to seven unimmunized heifers at the same stage of gestation. An additional unimmunized heifer was inoculated with uninfected monolayer cell culture material. All challenged heifers, immunized and unimmunized, had infected foetuses. Immunized heifers developed both parasite-specific humoral and cellular immune responses, characterised by increased IFAT titres, a predominant IgG1 response, elevated lymphoproliferative response and IFN-gamma production. Following tachyzoite challenge, they developed an anamnestic humoral response and produced similar amounts of IgG1 and IgG2 antibodies, but did not have an anamnestic cellular immune response. The lack of anamnestic cellular immune response and/or the large i.v/i.m tachyzoite inoculum may have contributed to the failure of the preparation.

Negligible prevalence of antibodies against Trypanosoma cruzi among blood donors in the southeastern United States.

Trypanosoma cruzi, a hemoflagellate, causes Chagas' disease and is endemic throughout Latin America. Increasing Latin American immigration to the United States has enhanced concern about transmission of Chagas' disease by infected donor blood. The insect vector and parasites also have been found in the southeastern United States. Autochthonous infection of several species of wild and domesticated mammals suggests that the general human population also may be at risk. To assess the prevalence of antibodies to T cruzi in humans, randomly selected donor blood was screened. Initial screening was performed by indirect hemagglutination (1:4 initial serum dilution) and at least one of three different enzyme immunoassays. All samples testing positive by at least one screening method were tested by radioimmunoprecipitation and indirect immunofluorescence supplemental methods, which were used for confirmation and calculation of specificity. Of the 6,013 serum samples evaluated, 85 tested positive by one screening method. Only 10 of the samples tested positive by more than one method. The percentages of positive screening tests are 0.05% by indirect hemagglutination and 0.06%, 0.91%, 3.97% by Abbott Laboratories (Abbott Park, Ill), Gull (Gull Laboratories, Salt Lake City, Utah), and Polychaco (Polychaco S.A.I.C., Buenos Aires, Argentina) enzyme immunoassays, respectively. All samples were negative by radioimmunoprecipitation and indirect immunofluorescence. These results suggest that although parasite and vector are found in the southeastern United States and both infect mammals, the risk of natural infection to humans in this region seems to be negligible. There was variation in positivity among different screening methods. The highest percentage of positive results was with the enzyme immunoassay, in which the binding of serum antibodies to antigens is amplified by enzymatic reactions.

Double-nested polymerase chain reaction for detection of caprine arthritis-encephalitis virus proviral DNA in blood, milk, and tissues of infected goats.

A nested polymerase chain reaction (PCR) for detecting proviral DNA of caprine arthritis-encephalitis virus (CAEV) in biological samples was developed. Primers for both gag and pol sequences of the CAEV genome were included in a single tube for simultaneous amplification ('double' PCR), and the resulting bands were resolved visually in ethidium bromide-stained agarose gels. Internal gag and pol probes were used to verify the identity of the amplified products by non-radioactive Southern hybridization. Final confirmation of the identity of representative PCR bands was provided by DNA sequence analysis. A comparison between the PCR and an antibody ELISA (with recombinant CAEV p28 as target) using 141 caprine blood samples indicated very strong agreement between the two assays (kappa = 0.912). Four of 7 goats with indeterminate ELISA results were PCR-positive as were 5 of 40 (12.5%) seronegative goats, most probably indicating delayed seroconversion. Eleven of 27 goats (41%) PCR-positive on blood had detectable CAEV proviral DNA in milk. Proviral DNA was also detected in lung, mesenteric lymph node, bone marrow, synovial membrane, and mammary gland of a seropositive, clinically affected goat, but not in equivalent tissues of a healthy seronegative goat.

Selective expression of a glutathione S-transferase subclass during spermatogenesis.

Levels of the hGSTM3 glutathione S-transferase (GST) subunit in testis of the human fetus and infant were found to be only a small fraction of those in adults. To understand these observations and to determine whether hGSTM3 subunit expression is developmentally and/or hormonally regulated, an experimental model based on the rat testis homologue (subunit rGSTM5) was used. For prepubertal rats, testicular rGSTM5 subunit levels were very low, but a sharp increase was observed between weeks 6 and 7 of development, when testicular growth includes increased numbers of germ cells associated with spermatogenesis. In adult hypophysectomized rats, the rGSTM5 subunit content of testis decreased progressively over 5 weeks, at which time the subunit was barely detectable. In contrast, the other GST subunit types did not vary significantly during development or after hypophysectomy. These results suggest that rGSTM5 subunits in rat testis could originate from spermatogenic cells. Accordingly, GSTs were purified from human sperm, and it was shown that the hGSTM3 subunit was, by a large measure, the predominant form. These data are consistent with the notion that the differential expression of hGSTM3 during human testicular development can also be explained on the basis of its preferential location in germs cells.

Human testicular glutathione S-transferases: insights into tissue-specific expression of the diverse subunit classes.

Cytosolic glutathione S-transferase (GST) subunits from human testis were resolved by HPLC and unambiguously identified by combined use of peptide sequence-specific antisera and electrospray ionization mass spectrometry (ESI MS). Allelic variants of hGSTP1, hGSTM1 and hGSTA2 were distinguished on the basis of observed differences in their molecular masses. Relative amounts of the multiple different subunit types in various human tissues were determined from HPLC profiles. From this type of analysis, tissues from hGSTM1 null allele individuals were readily discerned at the protein level; liver was the only tissue in which the hGSTM1 subunit was the major mu-class GST. hGSTM4 and hGSTM5 subunits were found at very low levels in all tissues examined. By far the tissue richest in the unique hGSTM3 subunit was testis, although brain also has significant levels.

A transient hypotensive episode (Bezold-Jarisch effect) occurring in a patient treated with microwave hyperthermia.

A case report of a hypotensive episode occurring during a microwave hyperthermia treatment is reported. Microwave hyperthermia was delivered using 915 MHz externally applied energy. The tumor being treated was either a superficial lymph node or local recurrence in the subdigastric region overlying the carotid artery. During the patient's treatment, she reported symptomatology consistent with a neuropathy which is characterized by its association with microwave energy. As the treatment progressed the patient developed syncopy, hypotension, and bradycardia similar to a Bezold-Jarisch reflex. All of these conditions promptly self-corrected when the microwave energy was discontinued. This event can be explained by a transient microwave induced neuropathy specifically involving the carotid body or sinus.

Fast neutron radiotherapy for soft tissue sarcomas. University of Washington experience and review of the world's literature.

Sixteen patients with inoperable soft tissue sarcomas were treated definitively with fast neutrons at the University of Washington between August, 1970 and May, 1982. Eleven of these 16 patients were treated with curative intent and form the basis of this report. Actuarial plots are shown for local tumor control and survival. This work is placed in the context of worldwide experience in using fast neutrons to treat unresectable soft tissue sarcomas.

Experimental reproduction of bovine fetal Neospora infection and death with a bovine Neospora isolate.

Studies were conducted to determine the pathogenic potential of the recently isolated bovine Neospora protozoa (BPA-1) for the bovine fetus. Cows chosen for study had Neospora titers < 160 using an indirect immunofluorescent antibody (IFA) test. Four experimental groups were studied. In group 1, 2 fetuses were inoculated in utero at 118 days gestation with culture-derived Neospora tachyzoites. A pregnant control cow was housed in the same pen, observed daily and screened serologically for evidence of exposure to Neospora. In group 2, 2 cows were infected with Neospora tachyzoites at 138 or 161 days gestation, and 1 control cow was given uninfected cell culture suspension simultaneously at 154 days gestation. Groups 3 (85 days gestation) and 4 (120 days gestation) each consisted of 2 cows infected with Neospora tachyzoites and 1 control cow given uninfected material at the same stage of gestation. Dead fetuses were surgically removed from the infected cows in group 1 on postinfection day (PID) 17. The histopathology was compatible with protozoal fetal infection, and protozoa were identified by immunohistochemistry. Viable fetuses were removed surgically from cows in group 2 on PID 28-30. The histopathology was compatible with protozoal fetal infection, protozoa were identified by immunoperoxidase techniques, and Neospora tachyzoites were reisolated in vitro from tissues of the 2 infected fetuses. In groups 3 and 4, the control fetus and 1 infected fetus were removed surgically between PID 26 and PID 33. The remaining infected cows were observed until fetal death or abortion occurred.(ABSTRACT TRUNCATED AT 250 WORDS)

Detection of Neospora sp. from infected bovine tissues by PCR and probe hybridization.

Neospora sp. can cause fetal abortion or neurological disease in congenitally infected calves. Latent tissue stages in infected cows may contribute to vertical transmission of Neospora sp. from dam to offspring in multiple pregnancies. In this investigation, the polymerase chain reaction (PCR) and Neospora-specific assay were employed to detect Neospora sp. by amplification of nuclear small-subunit rRNA gene sequences in infected cattle tissues. Tissues from 11 cattle, including 6 experimentally and 2 naturally infected cows, 1 naturally infected newborn calf, and 2 uninfected control cows, were evaluated in this study. Neospora-specific PCR products were amplified from DNAs of different bovine tissues, including brain, spinal cord, heart, lung, kidney, diaphragm, skeletal muscle, and placenta, as well as amniotic fluid samples of infected cattle. The PCR-based amplification and probe hybridization system proved useful in assessing the location of tissue-stage parasites in naturally and experimentally infected cattle, even when Neospora sp. antibody titers fall below normal cut-off values by an indirect immunofluorescent antibody test.

Successful pregnancy in goats carrying their genetically identical conceptus.

Mammalian pregnancies are naturally allogeneic, but syngeneic pregnancies have been carried to term in laboratory animal species. The need for maternal immune recognition during mammalian pregnancy is still unclear. Allogeneic pregnancies are protected from maternal immune attack by the nature of the trophoblast and its interactions with maternal tissues at the maternal-fetal interface. Syngeneic pregnancy models and the success of pregnancies in immunosuppressed mice challenge the necessity of a maternal immune response in mammals. This study was designed to investigate if outbred, domestic sheep and goats can successfully establish and maintain a syngeneic pregnancy. Embryo splitting and cryopreservation techniques were used to enable sheep and goat demi-embryos to be transferred to genetically identical females. Allogeneic pregnancies were established from the transfer of demi-embryos subjected to the same manipulations to assess demi-embryo survival and pregnancy rates under conventional immune compatibility conditions. Syngeneic pregnancies were established and carried to term in goats (2/11) but not in sheep (0/24). Microsatellite and DNA fingerprinting analyses confirmed that each kid was a genetically identical twin to the female that carried it to term. Our results demonstrated that genetic disparity is not required for the establishment and maintenance of pregnancy in goats, but our results were inconclusive for sheep.